1. Field of Invention
The present invention relates to a mutant prenyl diphosphate synthase that is able to synthesize prenyl diphosphate having a longer chain than prenyl diphosphate synthesized by the native prenyl diphosphate synthase.
2. Related Art
Prenyl diphosphate is highly valuable in biosynthesis pathways, functioning as a precursor of steroids, a precursor of carotenoids, being a transition substrate of prenylated proteins, being a substrate for synthesis of vitamin E, vitamin K and ubiquinone (CoQ) and so forth. Prenyl diphosphate exists in various forms, including dimethylallyl diphosphate (DMAPP; C5), geranyl diphosphate (GPP; C10), farnesyl diphosphate (FPP; C15), geranylgeranyl diphosphate (GGPP; C20), geranylfarnesyl diphosphate (GFPP; C25), hexaprenyl diphosphate (HPP; C30), heptaprenyl diphosphate (HepPP; C35) and octaprenyl diphosphate (OPP; C40).
Prenyl transferases, which synthesize these prenyl diphosphates, are enzymes that form prenyl diphosphate by continuously condensing isopentenyl diphosphate (IPP; C5) into allylic diphosphate, and exist in various forms, including farnesyl diphosphate synthase (FPS), geranylgeranyl diphosphate synthase (GGPS), geranylfarnesyl diphosphate synthase (GFPS), hexaprenyl diphosphate synthase (HexPS), heptaprenyl diphosphate synthase (HepPS) and octaprenyl diphosphate synthase (OPS).
However, among the above-mentioned prenyl diphosphates, only those from dimethylallyl diphosphate having 5 carbon atoms to geranyl diphosphate having 20 carbon atoms are commercially available in small amounts as reagents, and a process for industrially synthesizing and recovering large amounts of prenyl diphosphates having longer chains is not known.
The carbon chain length and stereoisomerism of synthesized prenyl diphosphates are known to be specifically determined depending on the particular enzyme. Until now, it has not been clear what type of mechanism is the factor in determining carbon chain length.
Although prenyl transferases and their genes are known to be derived from bacteria, mold, plants and animals, these enzyme are typically unstable, difficult to handle and are not expected to be industrially valuable.
The prenyl transferases and their genes of thermophilic organisms, which are stable and easy to use as enzymes, are only farnesyl diphosphate synthase (FPS) (Koyama, T. et al. (1995) J. Biol. Chem. 113, 355-363) and heptaprenyl diphosphate synthase (HepPS) (Koike-Takeshita, A. et al. (1995) J. Biol. Chem. 270, 18396-18400) from the moderately thermophilic archaebacterium, Bacillus stearothermophilus; geranylgeranyl diphosphate synthase (GGPS) from the hyper thermophilic bacterium, Sulfolobus acidocaldarius (Ohnuma, S.-i. et al. (1994) J. Biol. Chem. 268, 14792-14797); as well as farnesyl diphosphate/geranylgeranyl diphosphate synthase (FPS/GGPS) from the methane-producing archaebacterium, Methanobacterium thermoautotrophicum (Chen, A. and Poulter, C. D. (1993) J. Biol. Chem. 268, 11002-11007). Only HepPS can synthesize prenyl diphosphate having 35 carbon atoms, and enzymes having thermal stability that synthesize prenyl diphosphates having 25 or more carbon atoms have not been reported. In addition, the above-mentioned HepPS does not have adequate heat resistance, is composed of two types of subunits, and handling is not always easy.